The long-term goals of this proposal are to understand the physiologic regulation of phosphorylation of eIF-4E and the p25 subunit of eIF-4F and how this may effect translational control in vivo. Published studies have indicated that this protein is dephosphorylated in HeLa cells during the decrease in protein synthesis occurring after heat shock and during the mitotic phase of the cell cycle. Recent studies provide several lines of evidence for the existence of a novel nonabundant protein kinase in rabbit reticulocytes that phosphorylates eIF-4E in vitro at the same serine residue that in phosphorylated in intact reticulocytes and HeLa cells. The hypothesis to be tested is that a previously uncharacterized mammalian protein kinase (tentatively designated 4E kinase) exists which phosphorylates eIF-4E at serine-53 in vivo and that this phosphorylation event selectively modulates the translation of different mRNAs. In order to test this hypothesis the following immediate objectives are proposed: 1) purify the 4E kinase to enzymatic purity from rabbit reticulocytes using standard chromatography and HPLC methods and then characterize its substrate specificity and in vitro regulation; 2) prepare mouse monoclonal antibodies against the 4E kinase, immunohistochemical studies of cells and tissues to study the distribution of the enzyme and enzyme inhibition controls in cell-free translations and possibly intact cells; and 3) determine if 4E kinase has a mRNA-selective effect on translation in the reticulocyte lysate system. These studies may have implications for understanding the translational control mechanisms that occur in thermotolerance, the stress response and/or viral effects on host cells. Understanding the basic mechanisms of translational control may provide insight into the molecular mechanisms of diseases where a specific protein is synthesized in excess as in liver or pulmonary fibrosis.